Urticaria is a common skin disorder involving mast cell activation and degranulation. Urticaria is classified according to its chronicity into acute and chronic forms. It may occur spontaneously or upon exposure to a physical factor. In the latter case, the urticaria is classified as a physical urticaria. Physical urticaria may be induced by mechanical and applied pressure, exercise, or exposure to cold, heat, sun, water, or vibration. The pathologic basis of physical urticarias in general remains unclear and a genetic basis for these disorders is only partly understood. The purpose of this project is to investigate the mast cell dependent pathogenic mechanisms of physical urticaria, both to better understand how to manage urticarial inflammation and to explore the consequences of mast cell degranulation in human tissues. In these studies, patients undergo standard challenge testing to verify their urticaria. Blood samples are obtained for the investigation of molecular and genetic pathways involved in the disease process. Following the clinical induction of urticarial manifestations, additional blood samples are collected to determine soluble mediators involved in pathogenesis. Photographic imaging studies may be performed during challenge testing. Skin biopsies may be obtained prior to and following challenge testing and are analyzed for biochemical and histological markers. Since the inception of the physical urticaria protocol in 2009, we have enrolled over 154 patients including 25 healthy subjects. All patients safely underwent challenge testing based on their history. Blood samples and skin biopsies have been collected and stored for biochemical, molecular and, when applicable, genetic analysis. Mast cell degranulation was verified by skin biopsy. The majority of the patients were challenge positive to either cold-induced, cholinergic, dermatographism, solar or vibratory urticaria. Last year, we identified the genetic bases of vibratory urticaria based on findings from two large families, in which sustained vibration against the skin induces both a localized hive and systemic manifestations such as facial flushing. The findings of acute onset of symptom, and concurrent peripheral histamine release in affected family members implicated mast cell degranulation in the pathogenesis. Through whole exome sequencing we identified a missense mutation (p.C492Y) in the adhesion G-protein coupled receptor ADGRE2 that segregates with vibratory urticaria in affected individuals. ADGRE2 was determined to be highly expressed in human mast cells with a unique ligand ubiquitously found in the skin. This finding marked the first identification of a genetic basis for a mast cell-mediated urticaria induced by a mechanical stimulus and suggests that ADGRE2 subunit interactions may also have implications for other diseases involving mast cells (NEJM Feb 2016). Others affected with vibratory urticaria are being evaluated and screened for novel mutations in ADGRE2. Subsequent studies are underway in the MCBS to further elucidated the mechanism of ADGRE2 receptor activation and signaling in patients with inherited vibratory urticaria. In FY 2018, we continued a collaboration where serial samples from patients with cold urticaria are evaluated in a collaboration with Bruce Zuraw from UCSD to determine the activity of high molecular weight kinninogen (HMWK) as a measurement of bradykinin activity. Results thus far are unrevealing, as the levels of HMWK in patients were not different from controls. Additional indirect markers of activity are under investigation. In study continuing from FY 2017 into FY 2018, we did find that chromogranin A is not a useful marker to detect mast cell activation in patients with mast cell proliferative disorders. Elevated serum CgA levels were exclusively associated with proton pump inhibitor use, and did not correlate with mast cell burden or activation. In a challenge study in FY 2018, we documented an elevation in histamine and tryptase following exercise in patients with mastocytosis. In this study we investigated the effects of a treadmill exercise challenge on mast cell mediators in 8 patients diagnosed with indolent systemic mastocytosis (ISM) and compared the results to a patient with cholinergic urticaria and two normal volunteers. From venous serial serum samples at baseline and following exercise, we have reported that patients with ISM exhibit a significant and correlated rise in histamine and tryptase, consistent with the conclusion that physical activity can induce mast cell mediator release. Increase of these mediators in circulation was also associated with an exacerbation of mast cell mediator-related symptoms in the ISM cohort. This study in patients with ISM is consistent with an accumulating body of evidence that suggests physical exercise can trigger mast cell degranulation and that clinicians managing patients with ISM should provide instruction regarding the effects of exercise and guidance in the use of mast cell mediator receptor antagonists that may offer relief of symptoms, as well as the self-administration of epinephrine for the treatment of anaphylaxis should it occur.